Severe Hyperhomocysteinemia Promotes Bone Marrow-Derived and Resident Inflammatory Monocyte Differentiation and Atherosclerosis in LDLr/CBS-Deficient Mice

Rationale: Hyperhomocysteinemia (HHcy) accelerates atherosclerosis and increases inflammatory monocytes (MC) in peripheral tissues. However, its causative role in atherosclerosis is not well established and its effect on vascular inflammation has not been studied. The underlying mechanism is unknown. Objective: This study examined the causative role of HHcy in atherogenesis and its effect on inflammatory MC differentiation. Methods and Results: We generated a novel HHcy and hyperlipidemia mouse model, in which cystathionine beta-synthase (CBS) and low-density lipoprotein receptor (LDLr) genes were deficient (Ldlr(-/-) Cbs(-/-)). Severe HHcy (plasma homocysteine (Hcy)=275 mu mol/L) was induced by a high methionine diet containing sufficient basal levels of B vitamins. Plasma Hcy levels were lowered to 46 mu mol/L from 244 mu mol/L by vitamin supplementation, which elevated plasma folate levels. Bone marrow (BM)-derived cells were traced by the transplantation of BM cells from enhanced green fluorescent protein (EGFP) transgenic mice after sublethal irradiation of the recipient. HHcy accelerated atherosclerosis and promoted Ly6C(high) inflammatory MC differentiation of both BM and tissue origins in the aortas and peripheral tissues. It also elevated plasma levels of TNF-alpha, IL-6, and MCP-1; increased vessel wall MC accumulation; and increased macrophage maturation. Hcy-lowering therapy reversed HHcy-induced lesion formation, plasma cytokine increase, and blood and vessel inflammatory MC (Ly6C(high+middle)) accumulation. Plasma Hcy levels were positively correlated with plasma levels of proinflammatory cytokines. In primary mouse splenocytes, L-Hcy promoted rIFN gamma-induced inflammatory MC differentiation, as well as increased TNF-alpha, IL-6, and superoxide anion production in inflammatoryMCsubsets. Antioxidants and folic acid reversed L-Hcy-induced inflammatory MC differentiation and oxidative stress in inflammatory MC subsets. Conclusions: HHcy causes vessel wall inflammatory MC differentiation and macrophage maturation of both BM and tissue origins, leading to atherosclerosis via an oxidative stress-related mechanism. (Circ Res. 2012;111:37-49.)